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Colesterol , Receptores X del Hígado , Estado Prediabético , Transducción de Señal , Humanos , Estado Prediabético/metabolismo , Receptores X del Hígado/metabolismo , Receptores X del Hígado/genética , Colesterol/metabolismo , Receptores Citoplasmáticos y Nucleares/metabolismo , Receptores Citoplasmáticos y Nucleares/genética , Animales , Diabetes Mellitus/metabolismo , Factores de Riesgo , Diabetes Mellitus Tipo 2/metabolismo , RatonesRESUMEN
BACKGROUNDPreclinical studies suggest that cholesterol accumulation leads to insulin resistance. We previously reported that alterations in a monocyte cholesterol metabolism transcriptional network (CMTN) - suggestive of cellular cholesterol accumulation - were cross-sectionally associated with obesity and type 2 diabetes (T2D). Here, we sought to determine whether the CMTN alterations independently predict incident prediabetes/T2D risk, and correlate with cellular cholesterol accumulation.METHODSMonocyte mRNA expression of 11 CMTN genes was quantified among 934 Multi-Ethnic Study of Atherosclerosis (MESA) participants free of prediabetes/T2D; cellular cholesterol was measured in a subset of 24 monocyte samples.RESULTSDuring a median 6-year follow-up, lower expression of 3 highly correlated LXR target genes - ABCG1 and ABCA1 (cholesterol efflux) and MYLIP (cholesterol uptake suppression) - and not other CMTN genes, was significantly associated with higher risk of incident prediabetes/T2D. Lower expression of the LXR target genes correlated with higher cellular cholesterol levels (e.g., 47% of variance in cellular total cholesterol explained by ABCG1 expression). Further, adding the LXR target genes to overweight/obesity and other known predictors significantly improved prediction of incident prediabetes/T2D.CONCLUSIONThese data suggest that the aberrant LXR/ABCG1-ABCA1-MYLIP pathway (LAAMP) is a major T2D risk factor and support a potential role for aberrant LAAMP and cellular cholesterol accumulation in diabetogenesis.FUNDINGThe MESA Epigenomics and Transcriptomics Studies were funded by NIH grants 1R01HL101250, 1RF1AG054474, R01HL126477, R01DK101921, and R01HL135009. This work was supported by funding from NIDDK R01DK103531 and NHLBI R01HL119962.
Asunto(s)
Colesterol , Diabetes Mellitus Tipo 2 , Receptores X del Hígado , Estado Prediabético , Transducción de Señal , Humanos , Estado Prediabético/genética , Estado Prediabético/metabolismo , Masculino , Femenino , Diabetes Mellitus Tipo 2/genética , Diabetes Mellitus Tipo 2/metabolismo , Diabetes Mellitus Tipo 2/epidemiología , Persona de Mediana Edad , Receptores X del Hígado/genética , Receptores X del Hígado/metabolismo , Colesterol/metabolismo , Anciano , Transportador de Casetes de Unión a ATP, Subfamilia G, Miembro 1/genética , Transportador de Casetes de Unión a ATP, Subfamilia G, Miembro 1/metabolismo , Monocitos/metabolismo , Factores de Riesgo , Transportador 1 de Casete de Unión a ATP/genética , Transportador 1 de Casete de Unión a ATP/metabolismo , Anciano de 80 o más AñosRESUMEN
OBJECTIVE: The role of hepatocyte Abca1 (ATP binding cassette transporter A1) in trafficking hepatic free cholesterol (FC) into plasma versus bile for reverse cholesterol transport (RCT) is poorly understood. We hypothesized that hepatocyte Abca1 recycles plasma HDL-C (high-density lipoprotein cholesterol) taken up by the liver back into plasma, maintaining the plasma HDL-C pool, and decreasing HDL-mediated RCT into feces. Approach and Results: Chow-fed hepatocyte-specific Abca1 knockout (HSKO) and control mice were injected with human HDL radiolabeled with 125I-tyramine cellobiose (125I-TC; protein) and 3H-cholesteryl oleate (3H-CO). 125I-TC and 3H-CO plasma decay, plasma HDL 3H-CO selective clearance (ie, 3H-125I fractional catabolic rate), liver radiolabel uptake, and fecal 3H-sterol were significantly greater in HSKO versus control mice, supporting increased plasma HDL RCT. Twenty-four hours after 3H-CO-HDL injection, HSKO mice had reduced total hepatic 3H-FC (ie, 3H-CO hydrolyzed to 3H-FC in liver) resecretion into plasma, demonstrating Abca1 recycled HDL-derived hepatic 3H-FC back into plasma. Despite similar liver LDLr (low-density lipoprotein receptor) expression between genotypes, HSKO mice treated with LDLr-targeting versus control antisense oligonucleotide had slower plasma 3H-CO-HDL decay, reduced selective 3H-CO clearance, and decreased fecal 3H-sterol excretion that was indistinguishable from control mice. Increased RCT in HSKO mice was selective for 3H-CO-HDL, since macrophage RCT was similar between genotypes. CONCLUSIONS: Hepatocyte Abca1 deletion unmasks a novel and selective FC trafficking pathway that requires LDLr expression, accelerating plasma HDL-selective CE uptake by the liver and promoting HDL RCT into feces, consequently reducing HDL-derived hepatic FC recycling into plasma.
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Transportador 1 de Casete de Unión a ATP/fisiología , Colesterol/metabolismo , Hepatocitos/fisiología , Lipoproteínas HDL/sangre , Receptores de LDL/fisiología , Animales , Transporte Biológico , Femenino , Macrófagos/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BLRESUMEN
Although epidemiological data and results from rodent studies support an inverse relationship between nicotine consumption and body weight, the molecular mechanisms are poorly understood. CD-1 mice were fed a basal diet or a basal diet containing low or high dose smokeless tobacco blend or high dose nicotine tartrate for 14 weeks. High dose tobacco blend and nicotine tartrate diets vs. basal diet reduced mouse body weight (16.3% and 19.7%, respectively), epididymal (67.6% and 72.5%, respectively) and brown adipose weight (42% and 38%, respectively), epididymal adipocyte size (46.4% and 41.4%, respectively), and brown adipose tissue lipid droplet abundance, with no elevation of adipose tissue inflammation. High dose tobacco blend and nicotine diets also increased mouse physical activity and decreased respiratory exchange ratio, suggesting that high dose nicotine intake induces adipose tissue triglyceride lipolysis to provide fatty acids as an energy source. Both low and high dose tobacco blend and nicotine diet feeding vs. basal diet increased plasma insulin levels (2.9, 3.6 and 4.3-fold, respectively) and improved blood glucose disposal without affecting insulin sensitivity. Feeding of the high dose tobacco blend or nicotine feeding in mice induces body weight loss likely by increasing physical activity and stimulating adipose tissue triglyceride lipolysis.
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Adipocitos/efectos de los fármacos , Nicotiana , Nicotina/farmacología , Condicionamiento Físico Animal , Pérdida de Peso/efectos de los fármacos , Adipocitos/citología , Tejido Adiposo/metabolismo , Animales , Glucemia/metabolismo , Tamaño de la Célula , Conducta Alimentaria , Insulina/sangre , Resistencia a la Insulina , Lipólisis , Masculino , Ratones , Triglicéridos/metabolismoRESUMEN
OBJECTIVE: Adipose tissue cholesterol increases with adipocyte triglyceride content and size during development of obesity. However, how adipocyte cholesterol affects adipocyte function is poorly understood. The aim of this study was to evaluate the role of the cellular cholesterol exporter, Abca1 (ATP-binding cassette transporter A1), on adipose tissue function during diet-induced obesity. APPROACH AND RESULTS: Adiponectin Cre recombinase transgenic mice were crossed with Abca1flox/flox mice to generate ASKO (adipocyte-specific Abca1 knockout) mice. Control and ASKO mice were then fed a high-fat, high-cholesterol (45% calories as fat and 0.2% cholesterol) diet for 16 weeks. Compared with control mice, ASKO mice had a 2-fold increase in adipocyte plasma membrane cholesterol content and significantly lower body weight, epididymal fat pad weight, and adipocyte size. ASKO versus control adipose tissue had decreased PPARγ (peroxisome proliferator-activated receptor γ) and CCAAT/enhancer-binding protein expression, nuclear SREBP1 (sterol regulatory element-binding protein 1) protein, lipogenesis, and triglyceride accretion but similar Akt activation after acute insulin stimulation. Acute siRNA-mediated Abca1 silencing during 3T3L1 adipocyte differentiation reduced adipocyte Abca1 and PPARγ protein expression and triglyceride content. Systemic stimulated triglyceride lipolysis and glucose homeostasis were similar between control and ASKO mice. CONCLUSIONS: Adipocyte Abca1 is a key regulator of adipocyte lipogenesis and lipid accretion, likely because of increased adipose tissue membrane cholesterol, resulting in decreased activation of lipogenic transcription factors PPARγ and SREBP1.
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Transportador 1 de Casete de Unión a ATP/deficiencia , Adipocitos/metabolismo , Tejido Adiposo/metabolismo , Colesterol en la Dieta , Dieta Alta en Grasa , Eliminación de Gen , Lipólisis , Obesidad/prevención & control , Células 3T3-L1 , Transportador 1 de Casete de Unión a ATP/genética , Adipocitos/patología , Adipogénesis , Tejido Adiposo/patología , Tejido Adiposo/fisiopatología , Animales , Colesterol en la Dieta/metabolismo , Modelos Animales de Enfermedad , Predisposición Genética a la Enfermedad , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Obesidad/genética , Obesidad/metabolismo , Obesidad/fisiopatología , PPAR gamma/metabolismo , Fenotipo , Proteína 1 de Unión a los Elementos Reguladores de Esteroles/metabolismo , Triglicéridos/metabolismo , Aumento de PesoRESUMEN
G protein-coupled receptor (GPR)120/FFA receptor (FFAR)4 (GPR120/FFAR4) activation by n-3 PUFAs attenuates inflammation, but its impact on atherosclerosis is unknown. We determined whether in vivo activation of leukocyte GPR120/FFAR4 by n-3 versus n-6 PUFAs is atheroprotective. Leukocyte GPR120/FFAR4 WT or KO mice in the LDL receptor KO background were generated by bone marrow transplantation. Mice were fed one of the four atherogenic diets containing 0.2% cholesterol and 10% calories as palm oil (PO) + 10% calories as: 1) PO, 2) fish oil (FO; 20:5 n-3 and 22:6 n-3 enriched), 3) echium oil (EO; 18:4 n-3 enriched), or 4) borage oil (BO; 18:3 n-6 enriched) for 16 weeks. Compared with PO, mice fed BO, EO, and FO had significantly reduced plasma cholesterol, TG, VLDL cholesterol, hepatic neutral lipid, and atherosclerosis that were equivalent for WT and KO mice. In BO-, EO-, and FO-fed mice, but not PO-fed mice, lack of leukocyte GPR120/FFAR4 resulted in neutrophilia, pro-inflammatory Ly6Chi monocytosis, increased aortic root monocyte recruitment, and increased hepatic inflammatory gene expression. In conclusion, leukocyte GPR120 expression has minimal effects on dietary PUFA-induced plasma lipid/lipoprotein reduction and atheroprotection, and there is no distinction between n-3 versus n-6 PUFAs in activating anti-inflammatory effects of leukocyte GPR120/FFAR4 in vivo.
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Aterosclerosis/genética , Leucocitos/metabolismo , Receptores Acoplados a Proteínas G/genética , Receptores de LDL/genética , Animales , Aorta/metabolismo , Aorta/patología , Aterosclerosis/metabolismo , Aterosclerosis/patología , Colesterol/metabolismo , Dieta Aterogénica , Ácidos Grasos Omega-3/genética , Ácidos Grasos Omega-3/metabolismo , Ácidos Grasos Omega-6/genética , Ácidos Grasos Omega-6/metabolismo , Regulación de la Expresión Génica/efectos de los fármacos , Humanos , Inflamación/genética , Inflamación/metabolismo , Inflamación/patología , Leucocitos/patología , Ratones , Ratones Noqueados , Receptores Acoplados a Proteínas G/metabolismo , Activación Transcripcional/genéticaRESUMEN
OBJECTIVE: To test the hypothesis that the attenuation of cholesterol oleate packaging into apoB-containing lipoproteins will arrest progression of pre-existing atherosclerotic lesions. APPROACH AND RESULTS: Atherosclerosis was induced in apoB-100 only, LDLr(-/-) mice by feeding a diet enriched in cis-monounsaturated fatty acids for 24 weeks. A subset of mice was then euthanized to quantify the extent of atherosclerosis. The remaining mice were continued on the same diet (controls) or assigned to the following treatments for 16 weeks: (1) a diet enriched in n-3 polyunsaturated fatty acids, (2) the cis-monounsaturated fatty acid diet plus biweekly injections of an antisense oligonucleotide specific to hepatic sterol-O-acyltransferase 2 (SOAT2); or (3) the cis-monounsaturated fatty acid diet and biweekly injections of a nontargeting hepatic antisense oligonucleotide. Extent of atherosclerotic lesions in the aorta was monitored morphometrically in vivo with magnetic resonance imaging and ex vivo histologically and immunochemically. Hepatic knockdown of SOAT2 via antisense oligonucleotide treatment arrested lesion growth and stabilized lesions. CONCLUSIONS: Hepatic knockdown of SOAT2 in apoB100-only, LDLr(-/-) mice resulted in remodeling of aortic atherosclerotic lesions into a stable phenotype, suggesting SOAT2 is a viable target for the treatment of atherosclerosis.
Asunto(s)
Apolipoproteína B-100/sangre , ADN/genética , Regulación de la Expresión Génica , Hígado/enzimología , Oligonucleótidos Antisentido/genética , Placa Aterosclerótica/tratamiento farmacológico , Esterol O-Aciltransferasa/genética , Animales , Aorta Torácica/metabolismo , Aorta Torácica/patología , Modelos Animales de Enfermedad , Progresión de la Enfermedad , Imagen por Resonancia Magnética , Ratones , Ratones Noqueados , Oligonucleótidos Antisentido/farmacología , Placa Aterosclerótica/sangre , Placa Aterosclerótica/genética , Esterol O-Aciltransferasa/biosíntesis , Esterol O-Aciltransferasa/farmacología , Esterol O-Aciltransferasa 2RESUMEN
RATIONALE: Cholesterol esters (CE), especially cholesterol oleate, generated by hepatic and intestinal sterol O-acyltransferase 2 (SOAT2) play a critical role in cholesterol homeostasis. However, it is unknown whether the contribution of intestine-derived CE from SOAT2 would have similar effects in promoting atherosclerosis progression as for liver-derived CE. OBJECTIVE: To test whether, in low-density lipoprotein receptor null (LDLr(-/-)) mice, the conditional knockout of intestinal SOAT2 (SOAT2(SI-/SI-)) or hepatic SOAT2 (SOAT2(L-/L-)) would equally limit atherosclerosis development compared with the global deletion of SOAT2 (SOAT2(-/-)). METHODS AND RESULTS: SOAT2 conditional knockout mice were bred with LDLr(-/-) mice creating LDLr(-/-) mice with each of the specific SOAT2 gene deletions. All mice then were fed an atherogenic diet for 16 weeks. SOAT2(SI-/SI-)LDLr(-/-) and SOAT2(-/-)LDLr(-/-) mice had significantly lower levels of intestinal cholesterol absorption, more fecal sterol excretion, and lower biliary cholesterol levels. Analysis of plasma LDL showed that all mice with SOAT2 gene deletions had LDL CE with reduced percentages of cholesterol palmitate and cholesterol oleate. Each of the LDLr(-/-) mice with SOAT2 gene deletions had lower accumulations of total cholesterol and CE in the liver compared with control mice. Finally, aortic atherosclerosis development was significantly lower in all mice with global or tissue-restricted SOAT2 gene deletions. Nevertheless, SOAT2(-/-)LDLr(-/-) and SOAT2(L-/L-)LDLr(-/-) mice had less aortic CE accumulation and smaller aortic lesions than SOAT2(SI-/SI-)LDLr(-/-) mice. CONCLUSIONS: SOAT2-derived CE from both the intestine and liver significantly contribute to the development of atherosclerosis, although the CE from the hepatic enzyme appeared to promote more atherosclerosis development.
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Aorta/metabolismo , Aterosclerosis/metabolismo , Ésteres del Colesterol/metabolismo , Absorción Intestinal/fisiología , Hígado/metabolismo , Esterol O-Aciltransferasa/deficiencia , Animales , Aorta/patología , Aterosclerosis/sangre , Aterosclerosis/patología , Ésteres del Colesterol/sangre , Femenino , Masculino , Ratones , Ratones de la Cepa 129 , Ratones Endogámicos C57BL , Ratones Noqueados , Esterol O-Aciltransferasa 2RESUMEN
Several studies in humans and animals suggest that LDL particle core enrichment in cholesteryl oleate (CO) is associated with increased atherosclerosis. Diet enrichment with MUFAs enhances LDL CO content. Steroyl O-acyltransferase 2 (SOAT2) is the enzyme that catalyzes the synthesis of much of the CO found in LDL, and gene deletion of SOAT2 minimizes CO in LDL and protects against atherosclerosis. The purpose of this study was to test the hypothesis that the increased atherosclerosis associated with LDL core enrichment in CO results from an increased affinity of the LDL particle for arterial proteoglycans. ApoB-100-only Ldlr(-/-) mice with and without Soat2 gene deletions were fed diets enriched in either cis-MUFA or n-3 PUFA, and LDL particles were isolated. LDL:proteogylcan binding was measured using surface plasmon resonance. Particles with higher CO content consistently bound with higher affinity to human biglycan and the amount of binding was shown to be proportional to the extent of atherosclerosis of the LDL donor mice. The data strongly support the thesis that atherosclerosis was induced through enhanced proteoglycan binding of LDL resulting from LDL core CO enrichment.
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Aterosclerosis/metabolismo , Ésteres del Colesterol/metabolismo , LDL-Colesterol/metabolismo , Proteoglicanos/metabolismo , Resonancia por Plasmón de Superficie/métodos , Animales , Arterias/metabolismo , Biglicano/metabolismo , Humanos , RatonesRESUMEN
Reverse cholesterol transport (RCT) can proceed through the classic hepatobiliary route or through the nonbiliary transintestinal cholesterol efflux (TICE) pathway. Scavenger receptor class B type I (SR-BI) plays a critical role in the classic hepatobiliary route of RCT. However, the role of SR-BI in TICE has not been studied. To examine the role of intestinal SR-BI in TICE, sterol balance was measured in control mice and mice transgenically overexpressing SR-BI in the proximal small intestine (SR-BI(hApoCIII-ApoAIV-Tg)). SR-BI(hApoCIII-ApoAIV-Tg) mice had significantly lower plasma cholesterol levels compared with wild-type controls, yet SR-BI(hApoCIII-ApoAIV-Tg) mice had normal fractional cholesterol absorption and fecal neutral sterol excretion. Both in the absence or presence of ezetimibe, intestinal SR-BI overexpression had no impact on the amount of cholesterol excreted in the feces. To specifically study effects of intestinal SR-BI on TICE we crossed SR-BI(hApoCIII-ApoAIV-Tg) mice into a mouse model that preferentially utilized the TICE pathway for RCT (Niemann-Pick C1-like 1 liver transgenic), and likewise found no alterations in cholesterol absorption or fecal sterol excretion. Finally, mice lacking SR-BI in all tissues also exhibited normal cholesterol absorption and fecal cholesterol disposal. Collectively, these results suggest that SR-BI is not rate limiting for intestinal cholesterol absorption or for fecal neutral sterol loss through the TICE pathway.
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Colesterol/metabolismo , Absorción Intestinal/fisiología , Mucosa Intestinal/metabolismo , Receptores Depuradores de Clase B/metabolismo , Animales , Anticolesterolemiantes/farmacología , Azetidinas/farmacología , Colesterol/genética , Ezetimiba , Absorción Intestinal/efectos de los fármacos , Ratones , Ratones Transgénicos , Receptores Depuradores de Clase B/genéticaRESUMEN
Plant sterols, or phytosterols, are very similar in structure to cholesterol and are abundant in typical diets. The reason for poor absorption of plant sterols by the body is still unknown. Mutations in the ABC transporters G5 and G8 are known to cause an accumulation of plant sterols in blood and tissues (sitosterolemia). To determine the significance of phytosterol exclusion from the body, we fed wild-type and ABCG5/G8 knockout mice a diet enriched with plant sterols. The high-phytosterol diet was extremely toxic to the ABCG5/G8 knockout mice but had no adverse effects on wild-type mice. ABCG5/G8 knockout mice died prematurely and developed a phenotype that included high levels of plant sterols in many tissues, liver abnormalities, and severe cardiac lesions. This study is the first to report such toxic effects of phytosterol accumulation in ABCG5/G8 knockout mice. We believe these new data support the conclusion that plant sterols are excluded from the body because they are toxic when present at high levels.
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Transportadoras de Casetes de Unión a ATP/deficiencia , Conducta Alimentaria/efectos de los fármacos , Lipoproteínas/deficiencia , Fitosteroles/toxicidad , Transportador de Casetes de Unión a ATP, Subfamilia G, Miembro 5 , Transportador de Casete de Unión a ATP, Subfamilia G, Miembro 8 , Transportadoras de Casetes de Unión a ATP/metabolismo , Animales , Dieta , Eritrocitos/metabolismo , Regulación de la Expresión Génica/efectos de los fármacos , Hepatomegalia/sangre , Hepatomegalia/genética , Hepatomegalia/patología , Lipoproteínas/metabolismo , Hígado/efectos de los fármacos , Hígado/metabolismo , Hígado/patología , Ratones , Ratones Noqueados , Miocardio/metabolismo , Miocardio/patología , Tamaño de los Órganos/efectos de los fármacos , Bazo/efectos de los fármacos , Bazo/metabolismo , Bazo/patología , Esplenomegalia/sangre , Esplenomegalia/genética , Esplenomegalia/patología , Aumento de Peso/efectos de los fármacosRESUMEN
RATIONALE: Signal initiation by the high-density lipoprotein (HDL) receptor scavenger receptor class B, type I (SR-BI), which is important to actions of HDL on endothelium and other processes, requires cholesterol efflux and the C-terminal transmembrane domain. The C-terminal transmembrane domain uniquely interacts with plasma membrane (PM) cholesterol. OBJECTIVE: The molecular basis and functional significance of SR-BI interaction with PM cholesterol are unknown. We tested the hypotheses that the interaction is required for SR-BI signaling, and that it enables SR-BI to serve as a PM cholesterol sensor. METHODS AND RESULTS: In studies performed in COS-M6 cells, mutation of a highly conserved C-terminal transmembrane domain glutamine to alanine (SR-BI-Q445A) decreased PM cholesterol interaction with the receptor by 71% without altering HDL binding or cholesterol uptake or efflux, and it yielded a receptor incapable of HDL-induced signaling. Signaling prompted by cholesterol efflux to methyl-ß-cyclodextrin also was prevented, indicating that PM cholesterol interaction with the receptor enables it to serve as a PM cholesterol sensor. Using SR-BI-Q445A, we further demonstrated that PM cholesterol sensing by SR-BI does not influence SR-BI-mediated reverse cholesterol transport to the liver in mice. However, the PM cholesterol sensing does underlie apolipoprotein B intracellular trafficking in response to postprandial micelles or methyl-ß-cyclodextrin in cultured enterocytes, and it is required for HDL activation of endothelial NO synthase and migration in cultured endothelial cells and HDL-induced angiogenesis in vivo. CONCLUSIONS: Through interaction with PM cholesterol, SR-BI serves as a PM cholesterol sensor, and the resulting intracellular signaling governs processes in both enterocytes and endothelial cells.
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Membrana Celular/metabolismo , Colesterol/metabolismo , Células Endoteliales/metabolismo , Enterocitos/metabolismo , Receptores Depuradores de Clase B/metabolismo , Transducción de Señal , Alanina , Animales , Apolipoproteínas B/metabolismo , Células CACO-2 , Bovinos , Membrana Celular/efectos de los fármacos , HDL-Colesterol/metabolismo , Células Endoteliales/efectos de los fármacos , Enterocitos/efectos de los fármacos , Glutamina , Humanos , Péptidos y Proteínas de Señalización Intracelular/genética , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Hígado/metabolismo , Masculino , Proteínas de la Membrana , Ratones , Ratones Endogámicos C57BL , Mutación , Neovascularización Fisiológica , Óxido Nítrico Sintasa de Tipo III/metabolismo , Unión Proteica , Estructura Terciaria de Proteína , Receptores Depuradores de Clase B/química , Receptores Depuradores de Clase B/genética , Transducción de Señal/efectos de los fármacos , Factores de Tiempo , Transfección , beta-Ciclodextrinas/farmacologíaRESUMEN
The metabolic fate of newly absorbed cholesterol and phytosterol is orchestrated through adenosine triphosphate-binding cassette transporter G5 and G8 heterodimer (G5G8), and acyl CoA:cholesterol acyltransferase 2 (ACAT2). We hypothesized that intestinal G5G8 limits sterol absorption by reducing substrate availability for ACAT2 esterification and have attempted to define the roles of these two factors using gene deletion studies in mice. Male ACAT2(-/-), G5G8(-/-), ACAT2(-/-)G5G8(-/-) (DKO), and wild-type (WT) control mice were fed a diet with 20% of energy as palm oil and 0.2% (w/w) cholesterol. Sterol absorption efficiency was directly measured by monitoring the appearance of [(3)H]sitosterol and [(14)C]cholesterol tracers in lymph after thoracic lymph duct cannulation. The average percentage (± SEM) absorption of [(14)C]cholesterol after 8 h of lymph collection was 40.55 ± 0.76%, 19.41 ± 1.52%, 32.13 ± 1.60%, and 21.27 ± 1.35% for WT, ACAT2(-/-), G5G8(-/-), and DKO mice, respectively. [(3)H]sitosterol absorption was <2% in WT and ACAT2(-/-) mice, whereas it was up to 6.8% in G5G8(-/-) and DKO mice. G5G8(-/-) mice also produced chylomicrons with â¼70% less cholesterol ester mass than WT mice. In contrast to expectations, the data demonstrated that the absence of G5G8 led to decreased intestinal cholesterol esterification and reduced cholesterol transport efficiency. Intestinal G5G8 appeared to limit the absorption of phytosterols; ACAT2 more efficiently esterified cholesterol than phytosterols. The data indicate that handling of sterols by the intestine involves both G5G8 and ACAT2 but that an additional factor (possibly Niemann-Pick C1-like 1) may be key in determining absorption efficiency.
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Transportadoras de Casetes de Unión a ATP/metabolismo , Cateterismo , Lipoproteínas/metabolismo , Vasos Linfáticos/enzimología , Multimerización de Proteína , Esterol O-Aciltransferasa/metabolismo , Tórax , Transportador de Casetes de Unión a ATP, Subfamilia G, Miembro 5 , Transportador de Casete de Unión a ATP, Subfamilia G, Miembro 8 , Transportadoras de Casetes de Unión a ATP/química , Transportadoras de Casetes de Unión a ATP/deficiencia , Transportadoras de Casetes de Unión a ATP/genética , Animales , Colesterol/metabolismo , Esterificación , Regulación Enzimológica de la Expresión Génica , Técnicas de Inactivación de Genes , Absorción Intestinal , Lipoproteínas/química , Lipoproteínas/deficiencia , Lipoproteínas/genética , Masculino , Ratones , Estructura Cuaternaria de Proteína , Esterol O-Aciltransferasa/deficiencia , Esterol O-Aciltransferasa/genética , Esterol O-Aciltransferasa 2RESUMEN
Acyl-CoA:cholesterol acyltransferase 2 (ACAT2) generates cholesterol esters (CE) for packaging into newly synthesized lipoproteins and thus is a major determinant of blood cholesterol levels. ACAT2 is expressed exclusively in the small intestine and liver, but the relative contributions of ACAT2 expression in these tissues to systemic cholesterol metabolism is unknown. We investigated whether CE derived from the intestine or liver would differentially affect hepatic and plasma cholesterol homeostasis. We generated liver-specific (ACAT2(L-/L-)) and intestine-specific (ACAT2(SI-/SI-)) ACAT2 knockout mice and studied dietary cholesterol-induced hepatic lipid accumulation and hypercholesterolemia. ACAT2(SI-/SI-) mice, in contrast to ACAT2(L-/L-) mice, had blunted cholesterol absorption. However, specific deletion of ACAT2 in the intestine generated essentially a phenocopy of the conditional knockout of ACAT2 in the liver, with reduced levels of plasma very low-density lipoprotein and hepatic CE, yet hepatic-free cholesterol does not build up after high cholesterol intake. ACAT2(L-/L-) and ACAT2(SI-/SI-) mice were equally protected from diet-induced hepatic CE accumulation and hypercholesterolemia. These results suggest that inhibition of intestinal or hepatic ACAT2 improves atherogenic hyperlipidemia and limits hepatic CE accumulation in mice and that depletion of intestinal ACAT2 is sufficient for most of the beneficial effects on cholesterol metabolism. Inhibitors of ACAT2 targeting either tissue likely would be beneficial for atheroprotection.
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Colesterol/metabolismo , Dieta/efectos adversos , Técnicas de Inactivación de Genes , Mucosa Intestinal/metabolismo , Hígado/metabolismo , Esterol O-Aciltransferasa/deficiencia , Esterol O-Aciltransferasa/genética , Alelos , Animales , Sistema Biliar/metabolismo , Colesterol/sangre , Ésteres del Colesterol/metabolismo , Femenino , Hipercolesterolemia/etiología , Hipercolesterolemia/metabolismo , Hipercolesterolemia/prevención & control , Absorción Intestinal , Intestino Delgado/metabolismo , Ratones , Especificidad de Órganos , Esterol O-Aciltransferasa 2RESUMEN
The hypothesis tested in this study was that cholesterol esterification by ACAT2 would increase cholesterol absorption efficiency by providing cholesteryl ester (CE) for incorporation into chylomicrons. The assumption was that absorption would be proportional to Acat2 gene dosage. Male ACAT2âº/âº, ACAT2âº/â», and ACAT2â»/â» mice were fed a diet containing 20% of energy as palm oil with 0.2% (w/w) cholesterol. Cholesterol absorption efficiency was measured by fecal dual-isotope and thoracic lymph duct cannulation (TLDC) methods using [³H]sitosterol and [¹4C]cholesterol tracers. Excellent agreement among individual mice was found for cholesterol absorption measured by both techniques. Cholesterol absorption efficiency in ACAT2â»/â» mice was 16% compared with 46-47% in ACAT2âº/⺠and ACAT2âº/â» mice. Chylomicrons from ACAT2âº/⺠and ACAT2âº/â» mice carried â¼80% of total sterol mass as CE, whereas ACAT2â»/â» chylomicrons carried >90% of sterol mass in the unesterified form. The total percentage of chylomicron mass as CE was reduced from 12% in the presence of ACAT2 to â¼1% in ACAT2â»/â» mice. Altogether, the data demonstrate that ACAT2 increases cholesterol absorption efficiency by providing CE for chylomicron transport, but one copy of the Acat2 gene, providing â¼50% of ACAT2 mRNA and enzyme activity, was as effective as two copies in promoting cholesterol absorption.
Asunto(s)
Colesterol/metabolismo , Quilomicrones/metabolismo , Absorción Intestinal , Esterol O-Aciltransferasa/metabolismo , Animales , Ésteres del Colesterol/metabolismo , Esterificación , Masculino , Ratones , Tamaño de la Partícula , Conducto Torácico/metabolismo , Esterol O-Aciltransferasa 2RESUMEN
BACKGROUND: Adipose tissue (AT) is the body's largest free cholesterol reservoir and abundantly expresses ATP binding cassette transporter A1 (ABCA1), a key cholesterol transporter for high-density lipoprotein (HDL) biogenesis. However, the extent to which AT ABCA1 expression contributes to HDL biogenesis in vivo is unknown. METHODS AND RESULTS: Adipocyte-specific ABCA1 knockout mice (ABCA1(-A/-A)) were generated by crossing ABCA1(floxed) mice with aP2Cre transgenic mice. AT from ABCA1(-A/-A) mice had <10% of wild-type ABCA1 protein expression but normal hepatic and intestinal expression. Deletion of adipocyte ABCA1 resulted in a significant decrease in plasma HDL cholesterol (approximately 15%) and apolipoprotein A-I (approximately 13%) concentrations. AT from ABCA1(-A/-A) mice had a 2-fold increase in free cholesterol content compared with wild-type mice and failed to efflux cholesterol to apolipoprotein A-I. However, cholesterol efflux from AT to plasma HDL was similar for both genotypes of mice. Incubation of wild-type AT explants with apolipoprotein A-I resulted in the formation of multiple discrete-sized nascent HDL particles ranging in diameter from 7.1 to 12 nm; similar incubations with ABCA1(-A/-A) AT explants resulted in nascent HDL <8 nm. Plasma decay and tissue uptake of wild-type (125)I-HDL tracer were similar in both genotypes of recipient mice, suggesting that adipocyte ABCA1 deficiency reduces plasma HDL concentrations solely by reducing nascent HDL particle formation. CONCLUSIONS: We provide in vivo evidence that AT ABCA1-dependent cholesterol efflux and nascent HDL particle formation contribute to systemic HDL biogenesis and that AT ABCA1 expression plays an important role in adipocyte cholesterol homeostasis.
Asunto(s)
Transportadoras de Casetes de Unión a ATP/metabolismo , Tejido Adiposo/metabolismo , Lipoproteínas HDL/biosíntesis , Transportador 1 de Casete de Unión a ATP , Transportadoras de Casetes de Unión a ATP/genética , Adipocitos/metabolismo , Tejido Adiposo/citología , Animales , Apolipoproteína A-I/sangre , Colesterol/metabolismo , Homeostasis , Lipoproteínas HDL/sangre , Lipoproteínas HDL/química , Ratones , Ratones Noqueados , Ratones Transgénicos , Tamaño de la PartículaRESUMEN
Recent evidence suggests that the intestine may play a direct facilitative role in reverse cholesterol transport (RCT), independent of hepatobiliary secretion. In order to understand the nonbiliary pathway for RCT, we created both genetic and surgical models of biliary cholesterol insufficiency. To genetically inhibit biliary cholesterol secretion, we generated mice in which Niemann-Pick C1-Like 1 (NPC1L1) was overexpressed in the liver. Compared to controls, NPC1L1(Liver-Tg) mice exhibit a >90% decrease in biliary cholesterol secretion, yet mass fecal sterol loss and macrophage RCT are normal. To surgically inhibit biliary emptying into the intestine, we have established an acute biliary diversion model. Strikingly, macrophage RCT persists in mice surgically lacking the ability to secrete bile into the intestine. Collectively, these studies demonstrate that mass fecal sterol loss and macrophage RCT can proceed in the absence of biliary sterol secretion, challenging the obligate role of bile in RCT.
Asunto(s)
Sistema Biliar/metabolismo , Colesterol/metabolismo , Macrófagos/metabolismo , Esteroles/metabolismo , Animales , Transporte Biológico , Hígado/metabolismo , Macrófagos/inmunología , Proteínas de Transporte de Membrana/genética , Proteínas de Transporte de Membrana/metabolismo , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Modelos AnimalesRESUMEN
Acyl-CoA:cholesterol O-acyl transferase 2 (ACAT2) promotes cholesterol absorption by the intestine and the secretion of cholesteryl ester-enriched very low density lipoproteins by the liver. Paradoxically, mice lacking ACAT2 also exhibit mild hypertriglyceridemia. The present study addresses the unexpected role of ACAT2 in regulation of hepatic triglyceride (TG) metabolism. Mouse models of either complete genetic deficiency or pharmacological inhibition of ACAT2 were fed low fat diets containing various amounts of cholesterol to induce hepatic steatosis. Mice genetically lacking ACAT2 in both the intestine and the liver were dramatically protected against hepatic neutral lipid (TG and cholesteryl ester) accumulation, with the greatest differences occurring in situations where dietary cholesterol was elevated. Further studies demonstrated that liver-specific depletion of ACAT2 with antisense oligonucleotides prevents dietary cholesterol-associated hepatic steatosis both in an inbred mouse model of non-alcoholic fatty liver disease (SJL/J) and in a humanized hyperlipidemic mouse model (LDLr(-/-), apoB(100/100)). All mouse models of diminished ACAT2 function showed lowered hepatic triglyceride concentrations and higher plasma triglycerides secondary to increased hepatic secretion of TG into nascent very low density lipoproteins. This work demonstrates that inhibition of hepatic ACAT2 can prevent dietary cholesterol-driven hepatic steatosis in mice. These data provide the first evidence to suggest that ACAT2-specific inhibitors may hold unexpected therapeutic potential to treat both atherosclerosis and non-alcoholic fatty liver disease.
Asunto(s)
Hígado Graso/prevención & control , Hiperlipidemias/prevención & control , Hígado/metabolismo , Esterol O-Aciltransferasa/fisiología , Triglicéridos/metabolismo , Animales , Apolipoproteína B-100/fisiología , Western Blotting , Ésteres del Colesterol/metabolismo , Colesterol en la Dieta/administración & dosificación , Hígado Graso/metabolismo , Femenino , Hiperlipidemias/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Oligonucleótidos Antisentido/farmacología , ARN Mensajero/genética , ARN Mensajero/metabolismo , Receptores de LDL/fisiología , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Esterol O-Aciltransferasa/antagonistas & inhibidores , Esterol O-Aciltransferasa 2RESUMEN
Loss of ABCA1 activity in Tangier disease (TD) is associated with abnormal apoB lipoprotein (Lp) metabolism in addition to the complete absence of high density lipoprotein (HDL). We used hepatocyte-specific ABCA1 knock-out (HSKO) mice to test the hypothesis that hepatic ABCA1 plays dual roles in regulating Lp metabolism and nascent HDL formation. HSKO mice recapitulated the TD lipid phenotype with postprandial hypertriglyceridemia, markedly decreased LDL, and near absence of HDL. Triglyceride (TG) secretion was 2-fold higher in HSKO compared with wild type mice, primarily due to secretion of larger TG-enriched VLDL secondary to reduced hepatic phosphatidylinositol 3-kinase signaling. HSKO mice also displayed delayed clearance of postprandial TG and reduced post-heparin plasma lipolytic activity. In addition, hepatic LDLr expression and plasma LDL catabolism were increased 2-fold in HSKO compared with wild type mice. Last, adenoviral repletion of hepatic ABCA1 in HSKO mice normalized plasma VLDL TG and hepatic phosphatidylinositol 3-kinase signaling, with a partial recovery of HDL cholesterol levels, providing evidence that hepatic ABCA1 is involved in the reciprocal regulation of apoB Lp production and HDL formation. These findings suggest that altered apoB Lp metabolism in TD subjects may result from hepatic VLDL TG overproduction and increased hepatic LDLr expression and highlight hepatic ABCA1 as an important regulatory factor for apoB-containing Lp metabolism.
Asunto(s)
Transportadoras de Casetes de Unión a ATP/antagonistas & inhibidores , Transportadoras de Casetes de Unión a ATP/genética , Hepatocitos/metabolismo , Lipoproteínas LDL/metabolismo , Lipoproteínas VLDL/biosíntesis , Triglicéridos/biosíntesis , Transportador 1 de Casete de Unión a ATP , Transportadoras de Casetes de Unión a ATP/metabolismo , Adenoviridae/genética , Animales , Apolipoproteínas B/metabolismo , Modelos Animales de Enfermedad , Expresión Génica , Marcación de Gen , Humanos , Hipertrigliceridemia/genética , Hipertrigliceridemia/metabolismo , Lipoproteínas HDL/biosíntesis , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Fosfatidilinositol 3-Quinasas/metabolismo , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Enfermedad de Tangier/genética , Enfermedad de Tangier/metabolismoRESUMEN
BACKGROUND: Stearoyl-CoA desaturase 1 (SCD1) is a critical regulator of energy metabolism and inflammation. We have previously reported that inhibition of SCD1 in hyperlipidemic mice fed a saturated fatty acid (SFA)-enriched diet prevented development of the metabolic syndrome, yet surprisingly promoted severe atherosclerosis. In this study we tested whether dietary fish oil supplementation could prevent the accelerated atherosclerosis caused by SCD1 inhibition. METHODS AND RESULTS: LDLr(-/-), ApoB(100/100) mice were fed diets enriched in saturated fat or fish oil in conjunction with antisense oligonucleotide (ASO) treatment to inhibit SCD1. As previously reported, in SFA-fed mice, SCD1 inhibition dramatically protected against development of the metabolic syndrome, yet promoted atherosclerosis. In contrast, in mice fed fish oil, SCD1 inhibition did not result in augmented macrophage inflammatory response or severe atherosclerosis. In fact, the combined therapy of dietary fish oil and SCD1 ASO treatment effectively prevented both the metabolic syndrome and atherosclerosis. CONCLUSIONS: SCD1 ASO treatment in conjunction with dietary fish oil supplementation is an effective combination therapy to comprehensively combat the metabolic syndrome and atherosclerosis in mice.